Search

The alarming decline in sturgeon populations doubled by growing demands for sturgeon products, urge us to prevent the loss of these species. Fish stocking in natural habitats and developing fish farms are viable solutions, which can be successfully implemented with the help of reproductive biotechnologies. Despite the fact that semen cryopreservation is admittedly an important step for saving the Russian sturgeon, a reproducible standard method with good results has yet to be identified. Sperm quality assessment is essential for quantifying the impact of cryopreservation on spermatozoa. The purpose of our study was to provide additional information regarding semen cryopreservation and semen quality evaluation for the Russian sturgeon. Our study method is based on the use of two yolk-free extenders (with different cryoprotectants: DMSO and methanol) for freezing semen, using a simple freezing protocol. Parameters such as volume, concentration, motility, morphology and membrane integrity were evaluated. In conclusion, cryopreservation of Russian sturgeon spermatozoa using an extender containing methanol as cryoprotectant led to high egg fertilization percentages (72.67 ± 5.4%) even if the total motility values recorded for thawed semen were quite low (18-25%). Additionally, we identified two optimal stains for morphological studies and morphometry (Spermac stain kit and Trypan Blue Solution).

A severe, chronic and irreversible kidney disease affecting discrete rural populations in the Balkan Peninsula countries, Balkan endemic nephropathy (BEN) has been a scientific puzzle for more than half a century. Many environmental and other factors have been suggested as the primary cause and recent significant findings have linked BEN to aristolochic acids, phytotoxins derived from the plant Aristolochia clematitis, found in high density in the endemic areas. However, given that the incidence of BEN is less than 10% in affected villages, and it tends to have a family aggregation, as yet unidentified genetic factors may also play a role. To further explore this possibility, a pilot study was initiated to investigate the DNA methylation of CYP1A1, CYP1A2, NAT1, NQO1 and GSTT1 in blood samples from a group of Romanian BEN patients, compared to healthy controls and non-BEN chronic kidney disease (CKD) subjects. Our study revealed a more pronounced hypomethylation pattern in BEN and non-BEN CKD groups, compared to the healthy control group at specific CpGs across all five genes interrogated. Average methylation across the five regions investigated indicated significant differences only at GSTT1, in both BEN patients (p = 0.028) and non-BEN disease subjects (p = 0.015), relative to healthy individuals. Since GSTT1 active genotype appears to be a common feature of Serbian and Romanian BEN patients, GSTT1 epigenetic variation and increased gene activity could act as a predisposing (co)factor in BEN populations from the affected countries. BEN and non-BEN CKD groups show similar methylation patterns with exception of GSTT1 CpG8 (p = 0.046).

The aim of this study is to test the efficiency of different hormones on pikeperch spermiation and to establish a protocol for obtaining a high quantity of milt. Sixteen clinically healthy adult pikeperch males (3-4 years old) were used in our experiments. They were intraperitoneal injected with one of the following hormones: human chorionic gonadotropine - hCG (Pregnyl), GnRH analog (Receptal) and carp pituitary extract (CPE). These hormones were injected following two experimental protocols. The main differences between these two protocols are the number of stripings after the first injection (1 and 2 for the 1st protocol and the 2nd one, respectively) and the moment of thermal stimulation. The best results in stimulating spermiation were obtained with hCG and CPE. The protocols applicable for milt collecting in pikeperch males must include a thermal stimulation period.

The aim of our paper was to make a series of experiments in order to determine the concentration at which four cryoprotectants (DMSO, sucrose, Ficoll 70 and galactose) singly and in pairs would vitrify on plunging into liquid nitrogen and remain vitreous when thawed in water bath. As penetrating cryoprotector we used DMSO (MW=78.13 Da, Sigma D5879) and as nonpenetrating cryoprotectors we used sucrose (MW=342.3 Da, Sigma S7903), Ficoll 70 (MW= 60,000-80,000 Da, Sigma F4375) and Galactose (MW = 180,16 Da; Sigma G 6152). For DMSO there were tested concentrations from 1M to 6.5M, with concentrations step of 0.5M. For the nonpenetrating cryoprotectors there were tested concentrations of 5%, 10%, 15% and 20%. There were a total number of 168 solutions tested. The solutions vitrification ability on freezing was tested by direct plunging into liquid nitrogen at - 196°C. Three thawing temperatures were tested 20°C, 25°C and 37°C. The concentration at which DMSO solutions passed into vitreous state was 5M, but at thawing none of them remained vitreous at thawing. When pairs of cryoprotectors were tested 67 solutions vitrified at freezing (23 for DMSO-sucrose, 23 for DMSOFicoll 70 and 21 for DMSO-galactose) and 26 of them remained vitreous at thawing. The DMSO and galacose pair give the best results on thawing (11 solution remained vitreous on warming) at 37°C.

In order to evaluate the embryo viability we tested 2 cultivation media in three combinations (M16, Nutrient mixture F- 12 HAM supplemented with BSA and Nutrient mixture F- 12 HAM without BSA). In the third day of cultivation the embryos viability was assessed by morphological criteria and randomly by staining with fluorescents dyes (Fluorescein diacetate and Propidium iodide). It was observed that both media used for in vitro cultivation assured embryo development up to the expanded blastocyst stage (21.33% for M 16 and 4.34% respectively), exception the Nutrient mixture F- 12 HAM without BSA in which after 3 days cultivation all embryos were degenerated. The M16 medium assured the embryos development up to the hatched blastocyst stage (5.33%). None of the media supported the development of embryos under 8 cell stages and early blockage in stage of 2 cells could not be prevented.

The oocytes viability must be taken as an important selection parameter for successful in vitro cultivation. The ovaries were collected from the slaughterhouse and maintained at 4°C for 7 days. Fallowing cumulus - oocytes complexes recovery the viability was tested by two staining methods. For the first experiment we used 27 cumulus - oocytes complexes, stained with Neutral red and for the second experiment we used 11 cumulus - oocytes complexes stained with Trypan blue. Fallowing staining with Neutral red 23 cumulus - oocytes complexes were assessed as viable (were stained in red – enzymatic activity within the cells) and for the Trypan blue staining 11 cumulus - oocytes complexes were assessed as viable (remained unstained – integers cellular membranes).

The aim of the study was to determine if there is sexual dimorphism in a 2 years oldpikeperch population from the Danube. A total of 46 pikeperch (Sander lucioperca)individuals, males and females, caught in the Danube, were examined. Body weight,total length, standard length and girth as morphological traits and the Fulton indexwere calculated to compare males with females. The age of fish was determinedbased on the scales samples. Verge coefficient was taken into account whenclassifying individuals examined to age groups.Although there were some differences between males and females, the differenceswere not significant.

The effect of simple and sequential embryo culture media on the preimplantationdevelopment of mouse nuclear transfer (NT) embryos reconstructed with cumuluscell nuclei using a mechanical NT technique was studied. Blastocyst formation ratewas evaluated using CZB medium. Nonmanipulated and sham-manipulatedparthenogenetic embryos served as controls for, respectively, the medium and thehandling technique. Rates of blastocyst formation for medium and handling controlembryos were similar in CZB (50% and 53%). Development of NT embryos wassignificantly impaired from the two-cell stage onwards, reaching the blastocyst stage ata rate of 4% in CZB. These data demonstrate not only that NT embryos are moresensitive to in vitro culture conditions than parthenogenetic control embryos but alsothat selection of culture media can influence the preimplantation development of NTembryos.

Current researches were carried out with the goal to quantisize the lost from the weaning to early gestation at the sows housed in open pen gestation. In this trail we tested two pen types, different not only by size, but also by feeders’ emplacement. The main reproduction indicators that we calculated until the 28 gestation day were the proportion of sows in heat after weaning, the weaning to estrus interval and the gestation rates. The weaning to estrus interval was about 4 to 7 days, most sows were in heat in the day 5 and 6 days after weaning. The percent of heat detection after weaning was 71.42% for the small pens and 70.71% for the big pens (differences statistically non significant, chi test value was 0.983). The gestation rate at 28 days after insemination was 91.62% for the small pens and 94.72% for the large pens (chi test value 0,959, statistically non significant differences). The overpopulation for heat induction and after that chipping animals together in those pens, show that the lost are up to 40.47%, between weaning – day 28 of gestation.

Milk production and its composition are determined by quantitative loci, whichunder the influence of some environmental factors are producing an allelicvariability, meaning a genetic polymorphism of the gene. K-casein is a milk proteinwhose genetic polymorphism can serve as molecular marker for milk production,composition and industrial processing suitability. The allelic variants for k-casein Aand B are the most common and the most important of them. The experiments wereconducted on 24 Holstein-Friesian milking cows from a private farm in Giroc. Themilk production on a normal lactation is 8444 milk kg/305 days, with a fat percentof 3.9 and a protein percent of 3.3. The cows were divided in three groups AA, ABand BB in function of the genotyped obtained after the allelic variantsdetermination. The DNA isolation was made from hair roots and blood, the cowpopulation studied is not in genetic equilibrium fore k-casein gene, the frequency ofallele A is 0.43 and the frequency of B allele is 0.58. The highest genotype frequencywas 0.5 for CSN3-AB genotype, the BB genotype had 0.33 frequency, and the lowestfrequency was 0.17 for AA genotype. The mean daily milk production from cowswith BB genotype for k-casein is significant (p

During the experiments we have carried out with imature oocyte collected from the ovarian follicles, we found a variety of oocyte-cumulus complexes. We got the following experiment in order to understand the role of cumular cells on the achievement of the cytoplasma and oocyte nucleus maturation. We select the oocytecumulus complexes collected both from cows and sows according to the number of cumular cell layers and we watched their development to the blastocyst stade. Thus, we achieved three groups of COC (oocyte-cumulus complexes). One group was made of oocyte without cumular cells, the second group had a layer of cumular cells and the third group had many layers of cumular cells. we performed an incubation of all these types of COC in TCM-199 enriched with 20% of bovine fetal serum. Because only 1,2 oocyte of the ones who lack the cumular cells layer had maturation signs during cultivation in the thermostat versus 55 and 115, respectively, of the ones that had many cellular layers, presents a solid evidence that cumular cells are indispensable for the maturation and even to the fecundation process. The cumular cells perform a decisive role on the cytoplasma and oocyte nucleus maturation process.

The conception rate is defined as the aptitude of a female to romaine pregnant after the first A.I. and represents the proportion between the number of pregnant cows at first A.I and the number of cows inseminated in that period. In our experiments of estrus induction and synchronization, in order to increase the pregnancy rates, we used the Ovsynch protocol and other protocols for PgF2α and GnRH. In order to establish the pregnancy rates, the number of females that have not return in heats after A.I. were taken in consideration. The conception rates were 45.5 – 52.6%. The estrus synchronization offers the possibility to increase the pregnancy rates at first service after calving and creates the possibility to inseminate at certain time without heat detection.